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Abstract

Objective—To examine the impact of simple versus complex extracellular matrices (ECMs) on morphologic development and differentiation of bovine mammary gland progenitor cells (BMGPCs).

Sample Population—Cultures of BMGPCs.

Procedures—BMGPCs were grown on the following extracellular matrices: collagen I, collagen IV, laminin, and a commercially available gelatinous protein mixture. Cells were examined with light microscopy and transmission electron microscopy.

Results—Formation of organoids and production of the gap junction protein, connexin 43, were the criteria for BMGPC differentiation. The BMGPCs formed a 2-dimensional monolayer when grown on plastic, laminin, collagen I, or collagen IV. These cells did not have a network of cells forming epithelial organoids resembling a honeycomb. However, they did produce gap junction proteins. When BMGPCs were cultured on the commercially available gelatinous protein mixture, 3-dimensional epithelial organoids resembling a honeycomb formed and connexin 43 was produced. The thickness of the commercially available gelatinous protein mixture also regulated cell shape reorganization. Cell density affected the formation organoid networks and the rate at which monolayers reached confluency.

Conclusions and Clinical Relevance—When plated on a commercially available gelatinous protein mixture, the BMGPC culture system allowed us to simulate, in vitro, the interaction between epithelial cells in varying stages of differentiation and the microenvironment. Thus, a heterogenous ECM, such as the commercially available gelatinous protein mixture, is more physiologically relevant in providing a microenvironment for BMGPC lineage pathway differentiation to mimic an in vivo environment. In contrast, BMGPCs grown on homogenous ECM, although able to produce connexin 43, are unable to form organoids.

Full access
in American Journal of Veterinary Research
in Journal of the American Veterinary Medical Association

Abstract

Objective—To isolate bovine mammary gland cells with stem cell characteristics.

Sample Population—Monolayers of bovine mammary gland cells.

Procedure—Mammary gland cell populations were separated by use of selected media supplements. Phenotypic characteristics were examined via light and transmission electron microscopy. Cellular expression of casein and connexin 43 was identified immunohistochemically. A scrape-loading and dye transfer assay was used to examine the mammary gland cell populations for homogenous gap junctional intercellular communication (GJIC).

Results—Subpopulations of mammary gland cells grown in vitro are classified on the basis of their distinct morphologic features and ability to communicate via gap junctions. Ultrastructurally, 2 morphologically distinct cell types were classified as type I and II cells. Type I cells were small light undiffertiated cells and large light undifferentiated cells that were deficient in functional gap junctions (as is characteristic of stem cells). Type II cells included large light differentiated cells and terminally differentiated cells; GJIC was functional in type II cells. Type II cells had cytoplasmic expression of connexin 43, whereas, type I cells did not. All cells expressed casein.

Conclusions and Clinical Relevance—Subpopulations of bovine mammary gland cells with stem cell characteristics were identified. Phenotypic differences are observed among type I bovine mammary gland cells with stem cell characteristics. Gap junctional intercellular communication may be necessary for the differentiation of stem cells. Characterization of bovine mammary gland stem cells and their progeny may provide a new tool with which to study mammary gland health. (Am J Vet Res 2003;63:396–403)

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in American Journal of Veterinary Research

SUMMARY

Serum retinol, retinyl palmitate, and total vitamin A concentrations, and jejunoileal morphology were examined in neonatal calves infected with Cryptosporidium parvum. Group-1 calves served as noninfected controls and, after an adjustment period, were given 50 ml of saline solution iv every 12 hours for 6 days. Group-2 calves were inoculated with 107 C parvum oocysts and, after the onset of diarrhea, were given 50 ml of saline solution iv every 12 hours for 6 days. Group-3 calves were inoculated with 107 C parvum oocysts and, after the onset of diarrhea, were treated with difluoromethylornithine (dfmo, 200 mg/kg of body weight iv, q 12 h) for 6 days. Group-4 calves were naturally infected with C parvum. Jejunoileal biopsy specimens were excised from calves of groups 1-3 at 3 and again at 15 to 16 days of age. During the course of diarrhea and 3 days after saline or dfmo administration, water-miscible retinyl palmitate was administered orally (2,750 μg/kg) to each calf in each group. Cryptosporidium parvum infection was associated with significant (P ≤ 0.05) reduction in postadministration serum retinol, retinyl palmitate, and total vitamin A concentrations in calves of groups 2, 3, and 4. Cryptosporidium parvum infection caused significant (P ≤ 0.05) reduction in villus height. Decreased villus height, villus blunting and fusion, and attenuation of the intestinal mucosa were associated with reduced absorption of vitamin A, as indicated by lower peak postadministration retinyl palmitate concentration in C parvum-infected calves. Intravenous administration of dfmo to group-3 calves did not improve retinol absorption. Vitamin A should be provided parenterally to young calves with enteric cryptosporidiosis in an attempt to avoid depletion of concurrent low liver vitamin A reserves.

Free access
in American Journal of Veterinary Research

Abstract

Objective

To determine pharmacokinetics of single and multiple doses of rimantadine hydrochloride in horses and to evaluate prophylactic efficacy of rimantadine in influenza virus-infected horses.

Animals

5 clinically normal horses and 8 horses seronegative to influenza A.

Procedure

Horses were given rimantadine (7 mg/kg of body weight, IV, once; 15 mg/kg, PO, once; 30 mg/kg, PO, once; and 30 mg/kg, PO, q 12 h for 4 days) to determine disposition kinetics. Efficacy in induced infections was determined in horses seronegative to influenza virus A2. Rimantadine was administered (30 mg/kg, PO, q 12 h for 7 days) beginning 12 hours before challenge-exposure to the virus.

Results

Estimated mean peak plasma concentration of rimantadine after IV administration was 2.0 µg/ml, volume of distribution (mean ± SD) at steady-state (VdSS) was 7.1 ± 1.7 L/kg, plasma clearance after IV administration was 51 ± 7 ml/min/kg, and β-phase halflife was 2.0 ± 0.4 hours. Oral administration of 15 mg of rimantadine/kg yielded peak plasma concentrations of < 50 ng/ml after 3 hours; a single oral administration of 30 mg/kg yielded mean peak plasma concentrations of 500 ng/ml with mean bioavailability (F) of 25%, β-phase half-life of 2.2 ± 0.3 hours, and clearance of 340 ± 255 ml/min/kg. Multiple doses of rimantadine provided steady-state concentrations in plasma with peak and trough concentrations (mean ± SEM) of 811 ± 97 and 161 ± 12 ng/ml, respectively. Rimantadine used prophylactically for induced influenza virus A2 infection was associated with significant decreases in rectal temperature and lung sounds.

Conclusions and Clinical Relevance

Oral administration of rimantadine to horses can safely ameliorate clinical signs of influenza virus infection. (Am J Vet Med 1999:60:888–894)

Free access
in American Journal of Veterinary Research

Summary:

Eight of 19 calves born to bovine viral diarrhea virus (bvdv)-negative and -immunocompetent dams were determined to be infected with a noncytopathic strain of bvdv. Six of the 8 calves had diarrhea and 2 had no clinical signs of disease. In 3 euthanatized calves, lesions consistent with mucosal disease were found throughout the gastrointestinal tract, and the virus was isolated from the spleen, lymph nodes, and small intestine. In 5 calves, bvdv was isolated from mononuclear cells in blood samples obtained 21 days apart, indicating persistent infection. The virus was not isolated from sera obtained from 2 calves, with chronic nonclinical infections, that had neutralizing antibody titers ≥ 1:512 against bovine viral diarrhea-Singer virus and titers ≥ 1:256 against the persistent bvdv. Twenty-one days after vaccination with a vaccine that contained inactivated noncytopathic and cytopathic biotypes of bvdv, 4 of 5 persistently infected calves had neutralizing antibody titers ≤ 1:4 against the bovine viral diarrhea-Singer virus and their persistent virus. Prior to vaccination, 2 of 11 virus-negative calves had neutralizing antibody titers ≤ 1:128 against the bovine viral diarrhea-Singer virus, and after vaccination, only 1 virus-negative calf had a titer ≤ 1:512. At 149 days after revaccination and prior to weaning, 4 virus-negative calves had neutralizing antibody titers ≤ 1:512 (range, 1:16 to 1:384). Under the specific conditions in this herd, we were not able to detect a beneficial effect of vaccination. Colostral origin bvdv-specific antibody, capable of neutralizing the persistently infective bvdv strain, most likely interfered with isolation of the virus from the sera of 2 persistently infected calves.

Free access
in Journal of the American Veterinary Medical Association

Objective—

To determine gross income lost that was attributable to thin cows in a beef cattle herd, to estimate the cost of added nutrition necessary to prevent thin cows in the herd, and to determine the financial outcome of the improved nutritional practices.

Design—

Prospective, observational study.

Animals—

Four hundred twenty-two Santa Gertrudis cows and their calves.

Procedure—

At pregnancy examination in the fall of 1992, cows were assigned a body condition score (BCS), using a scale of 1 (emaciated) to 9 (obese), and the ratio of the productivity of BCS-3 and BCS-4 cow groups (thin cows), compared with the mean productivity of BCS-5 and BCS-6 cows (cows in good condition), was determined. Measures of productivity evaluated included pregnancy rates, weaning weights, and prices per hundredweight of calves. The performance ratios of BCS-3 and BCS-4 cows were multiplied by the mean gross income of BCS-5 and BCS-6 cows to calculate their gross income. This was then subtracted from the mean income of BCS-5 and BCS-6 cows to estimate the amount of lost gross income per thin cow. The cost of a nutritional program that would prevent thin cows in the herd was subtracted from the lost gross income of the thin cows to yield the amount of increased net income that could be generated from a nutritional program that would maintain cows in the herd at a BCS of 5 or 6.

Results—

Cows with a BCS of 3 were 0.48 as productive, and cows with a BCS of 4 were 0.74 as productive as the average of the BCS-5 and BCS-6 cows combined. Each BCS-3 cow generated $215.06 less, and each BCS-4 cow generated $107.53 less gross income than the average gross income of BCS-5 and BCS-6 cows. The added cost of nutrition that would have reconditioned BCS-3 and BCS-4 cows to a BCS of 5.5 was $91.48/BCS-3 cow and $43.67/ BCS-4 cow. Implementation of the reconditioning nutrition program the previous fall would have resulted in an extra net income of $123.58/BCS-3 cow and $63.86/BCS-4 cow, received over a 2-year period. The 262 thin cows in the herd accounted for a total net income loss of $19,897.

Clinical Implications—

The time of pregnancy examination is a strategic intervention point to estimate the past negative economic impact of thin cows and to implement a plan to prevent these losses in the future.

Free access
in Journal of the American Veterinary Medical Association

Abstract

Objective—To determine safety, efficacy, and immunogenicity of an intranasal cold-adapted modified- live equine influenza virus vaccine administered to ponies following induction of exercise-induced immunosuppression.

Design—Prospective study.

Animals—Fifteen 9- to 15-month old ponies that had not had influenza.

Procedure—Five ponies were vaccinated after 5 days of strenuous exercise on a high-speed treadmill, 5 were vaccinated without undergoing exercise, and 5 were not vaccinated or exercised and served as controls. Three months later, all ponies were challenged by nebulization of homologous equine influenza virus. Clinical and hematologic responses and viral shedding were monitored, and serum and nasal secretions were collected for determination of influenza-virus-specific antibody isotype responses.

Results—Exercise caused immunosuppression, as indicated by depression of lymphocyte proliferation in response to pokeweed mitogen. Vaccination did not result in adverse clinical effects, and none of the vaccinated ponies developed clinical signs of infection following challenge exposure. In contrast, challenge exposure caused marked clinical signs of respiratory tract disease in 4 control ponies. Vaccinated and control ponies shed virus after challenge exposure. Antibody responses to vaccination were restricted to serum IgGa and IgGb responses in both vaccination groups. After challenge exposure, ponies in all groups generated serum IgGa and IgGb and nasal IgA responses. Patterns of serum hemagglutination inhibition titers were similar to patterns of IgGa and IgGb responses.

Conclusions and Clinical Relevance—Results suggested that administration of this MLV vaccine to ponies with exercise-induced immunosuppression was safe and that administration of a single dose to ponies provided clinical protection 3 months later. (J Am Vet Med Assoc 2001;218:900–906)

Full access
in Journal of the American Veterinary Medical Association

Abstract

Objective—To evaluate serum haptoglobin concentration at feedlot arrival and subsequent performance and morbidity and mortality rates of calves that developed bovine respiratory disease.

Animals—360 heifer calves and 416 steer and bull calves.

Procedures—Serum samples were obtained from cattle at the time of arrival to a feedlot (day −1) and analyzed for haptoglobin concentration. In experiment 1, calves were classified into groups with a low (< 1.0 μg/mL), medium (1.0 to 3.0 μg/mL), or high (> 3.0 μg/mL) serum haptoglobin concentration and allotted into pens on the basis of group. In experiment 2, calves were classified as having or not having detectable serum haptoglobin concentrations.

Results—In experiment 1, average daily gain from days 1 to 7 decreased as haptoglobin concentration increased. Dry-matter intake (DMI) from days 1 to 21 decreased with increasing haptoglobin concentration, and DMI typically decreased from days 1 to 63. Total bovine respiratory disease morbidity rate typically increased with increasing haptoglobin concentration. At harvest, no differences in carcass characteristics were observed on the basis of haptoglobin concentration. In experiment 2, cattle with measureable serum haptoglobin concentrations at arrival weighed less throughout the experiment, gained less from days 1 to 7, and had lower DMI from days 1 to 42. Overall morbidity rate was not different between groups, but cattle with detectable serum haptoglobin concentrations had higher odds of being treated 3 times.

Conclusions and Clinical Relevance—Serum haptoglobin concentration in cattle at the time of feedlot arrival was not associated with overall performance but may have limited merit for making decisions regarding targeted prophylactic treatment.

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in American Journal of Veterinary Research

Abstract

Objective—To determine efficacy of a modified-live virus (MLV) vaccine containing bovine viral diarrhea virus (BVDV) 1a and 2a against fetal infection in heifers exposed to cattle persistently infected (PI) with BVDV subtype 1 b.

Animals—50 heifers and their fetuses.

Procedures—Susceptible heifers received a placebo vaccine administered IM or a vaccine containing MLV strains of BVDV1a and BVDV2a administered IM or SC. On day 124 (64 to 89 days of gestation), 50 pregnant heifers (20 vaccinated SC, 20 vaccinated IM, and 10 control heifers) were challenge exposed to 8 PI cattle. On days 207 to 209, fetuses were recovered from heifers and used for testing.

Results—2 control heifers aborted following challenge exposure; both fetuses were unavailable for testing. Eleven fetuses (8 control heifers and 1 IM and 2 SC vaccinates) were positive for BVDV via virus isolation (VI) and for BVDV antigen via immunohistochemical analysis in multiple tissues. Two additional fetuses from IM vaccinates were considered exposed to BVDV (one was seropositive for BVDV and the second was positive via VI in fetal tissues). A third fetus in the SC vaccinates was positive for BVDV via VI from serum alone. Vaccination against BVDV provided fetal protection in IM vaccinated (17/20) and SC vaccinated (17/20) heifers, but all control heifers (10/10) were considered infected.

Conclusions and Clinical Relevance—1 dose of a BVDV1a and 2a MLV vaccine administered SC or IM prior to breeding helped protect against fetal infection in pregnant heifers exposed to cattle PI with BVDV1b.

Full access
in American Journal of Veterinary Research